Différenciation des cellules du lignage antéhypophysaire somatolactotrope (un r^ole pour les facteurs de transcription Pitx1 et Pitx2)

AIX-MARSEILLE2-BU Sci.Luminy (130552106) / SudocSudocFranceF

Truncation of PITX2 differentially affects its activity on physiological targets.

International audienceThe bicoid-like transcription factor PITX2 has been previously described to interact with the pituitary-specific POU homeodomain factor POU1F1 (human ortholog of PIT-1) to achieve cell-specific expression of prolactin (PRL) and GH in pituitary somatolactotroph cells. In this work, we have investigated the functional properties of three PITX2 mutants reported in Axenfeld-Rieger syndrome patients relative to the regulation of these genes, using reporter genes under the control of human PRL (hPRL), hGH, or POU1F1 promoters transfected in nonpituitary and pituitary cell lines. Among the three mutations studied, Y167X and E101X introduce a premature stop codon, and F104L leads to an amino acid substitution. While PITX2(E101X) is not expressed in the cells following transfection, and PITX2(F104L) is functionally inactive, the PITX2(Y167X) mutant keeps its DNA-binding capacity and displays a markedly enhanced activation of the hPRL and POU1F1 promoters, but not of the hGH promoter. Y167X is the first mutation of PITX2 described to result in a differential effect on the activation of its different physiological targets, hPRL and POU1F1 on one hand and hGH on the other hand. The differential effect of the Y167X mutation might be linked to an interaction of PITX2 with different transcription factors or cofactors when bound to the hPRL and POU1F1 or the hGH promoters. These results might form the basis for the identification of the PITX2 protein complex necessary for the differential GH or PRL expression

Heterozygous LHX3 mutations may lead to a mild phenotype of combined pituitary hormone deficiency

International audienceLHX3 is an LIM domain transcription factor involved in the early steps of pituitary ontogenesis. We report here functional studies of three allelic variants, including the first heterozygous variant of LHX3 NM_178138.5(LHX3)c.587T>C (p.(Leu196Pro)) that may be responsible for a milder phenotype of hypopituitarism. Our functional studies showed that NM_178138.5(LHX3)c.587T>C (p.(Leu196Pro)) was not able to activate target promoters in vitro, as it did not bind DNA, and likely affected LHX3 function via a mechanism of haplo-insufficiency. Our study demonstrates the possibility that patients with a heterozygous variant of LHX3 may have pituitary deficiencies, with a milder phenotype than patients with homozygous variants. It is thus of vital to propose an optimal follow-up of such patients, who, until now, were considered as not being at risk of presenting pituitary deficiency. The second variant NM_178138.5(LHX3)c.622C>G (p.(Arg208Gly)), present in a homozygous state, displayed decreased transactivating ability without loss of binding capacity in vitro, concordant with in silico analysis; it should thus be considered to affect LHX3 function. In contrast, the NM_178138.5(LHX3)c.929G>C (p.(Arg310Pro)) variant, in a heterozygous state, also predicted as deleterious in silico, proved functionally active in vitro, and should thus still be classified as a variant of unknown significance. Our study emphasizes the need for functional studies due to the limits of software-based predictions of new variants, and the possible association of a pituitary phenotype to heterozygous LHX3 variants

Mutations in NFKB2 and potential genetic heterogeneity in patients with DAVID syndrome, having variable endocrine and immune deficiencies.

International audienceDAVID syndrome is a rare condition combining anterior pituitary hormone deficiency with common variable immunodeficiency. NFKB2 mutations have recently been identified in patients with ACTH and variable immunodeficiency. A similar mutation was previously found in Nfkb2 in the immunodeficient Lym1 mouse strain, but the effect of the mutation on endocrine function was not evaluated. We ascertained six unrelated DAVID syndrome families. We performed whole exome and traditional Sanger sequencing to search for causal genes. Lym1 mice were examined for endocrine developmental anomalies. Mutations in the NFKB2 gene were identified in three of our families through whole exome sequencing, and in a fourth by direct Sanger sequencing. De novo origin of the mutations could be demonstrated in three of the families. All mutations lie near the C-terminus of the protein-coding region, near signals required for processing of NFΚB2 protein by the alternative pathway. Two of the probands had anatomical pituitary anomalies, and one had growth and thyroid hormone as well as ACTH deficiency; these findings have not been previously reported. Two children of one of the probands carried the mutation and have to date exhibited only an immune phenotype. No mutations were found near the C-terminus of NFKB2 in the remaining two probands; whole exome sequencing has been performed for one of these. Lym1 mice, carrying a similar Nfkb2 C-terminal mutation, showed normal pituitary anatomy and expression of proopiomelanocortin (POMC). We confirm previous findings that mutations near the C-terminus of NFKB2 cause combined endocrine and immunodeficiencies. De novo status of the mutations was confirmed in all cases for which both parents were available. The mutations are consistent with a dominant gain-of-function effect, generating an unprocessed NFKB2 super-repressor protein. We expand the potential phenotype of such NFKB2 mutations to include additional pituitary hormone deficiencies as well as anatomical pituitary anomalies. The lack of an observable endocrine phenotype in Lym1 mice suggests that the endocrine component of DAVID syndrome is either not due to a direct role of NFKB pathways on pituitary development, or else that human and mouse pituitary development differ in its requirements for NFKB pathway function